1. Leptin
Leptin is produced from the obese (ob) gene and binds to the ob receptors (Ob-R). The ob gene is expressed in various tissues such as placenta, ovaries, muscle and adipose tissue. Leptin is produced in the adipocyte and in ovaries, and is a circulating 16 kDa protein (G. A. Bray, (1996) Lancet 348: 140; C. Liu et al., (1997) Endocrinology 138: 3548). Defective production of leptin results in gross obesity and type 2 diabetes in the obese (ob/ob) mouse. In humans, the leptin protein levels have been correlated to the percentage of body fat and is elevated in obese patients (R. V. Considine et al., (1996) N. Engl. J. Med. 334: 292). Defects in the leptin receptor, Ob-Rb, produces a syndrome in the mutant diabetic db/db mouse that is phenotypically identical to that observed in the ob/ob mouse. In addition to obesity, leptin is also believed to modulate estrogen expression and the fat stores needed for reproduction purposes. Other potential roles for leptin include regulation of hemopoiesis and macrophage function (T. Gainforth et al., (1996) Proc. Nat'l Acad. Sci. USA 93: 14564).
Leptin has been detected in the plasma of normal individuals and individuals receiving hemodialysis and in renal transplant patients, in placental tissue from pregnant women, and in cord blood of newborns (Respectively, J. K. Howard et al., (1997) Clin. Sci. 93: 119; S. G. Hassink et al., (1997) Pediatrics 100: 123). It has been suggested that leptin concentrations in newborns cannot be explained by adiposity alone. In women, leptin deficiency has been postulated to be involved with delayed puberty, menstrual disturbances and anorexia nervosa (M. Schwartz et al., (1997) N. Engl. J. Med. 336: 1802). Leptin is also believed to regulate lipid metabolism, glucose uptake, β-cell function, gonadotropin secretion, sympathetic tone, ovarian function and thermogenesis. Glucocorticoids and insulin increase leptin production. Administration of leptin reduces food intake, decreases insulin concentrations, and lowers blood glucose concentrations in the ob/ob mouse, but not in the db/db mouse (G. A. Bray, (1996) Lancet 348: 140).
2. The Leptin Receptor
The leptin receptor belongs to the cytokine superfamily of receptors. Several forms of the leptin receptor are expressed in humans and rodents (G. A. Bray, (1996) Lancet 348: 140). The short form (Ob-R(S)), considered to have limited signaling capability, is detected in many organs and has 5 identified isoforms, Ob-Ra, Ob-Rc, Ob-Rd, Ob-Re, and r-Ob-Rf (M.-Y. Wang et al., (1996) FEBS Letters 392: 87). Ob-R(S) has been identified in the choroid plexus and may be involved in the transport of leptin across the blood-brain barrier (J. Girard, (1997) Diabetes Metabol. 23S: 16).
It is the long form of the leptin receptor which is believed to mediate the biological effects of the leptin protein (L. A. Campfield et al., (1996) Horm. Metab. Res. 28: 619). In contrast to the short form of the leptin receptor, Ob-R long form (Ob-R(L) also known as Ob-Rb) predominates in the hypothalamus and cerebellum (A. Savioz et al., (1997) Neuroreport 8: 3123; J. G. Mercer et al., (1996) FEBS Letters 387: 113). Ob-R(L) has also been detected at low concentrations in peripheral tissues (Y. Wang et al., (1997) J. Biol. Chem. 272: 16216), such as the brain (A. Heritier et al., (1997) Neurosci. Res. Commun. 21: 113), spleen, testes, kidney, liver, lung, adrenal (N. Hoggard et al., (1997) Biochem. Biophys. Res. Commun. 232: 383), and hematopoietic tissues (A. A. Mikhail et al., (1997) Blood 89: 1507). Ob-R(L) has also been observed in the placenta, fetal cartilage/bone, and hair follicles, and therefore is believed to play a role in development (N. Hoggard et al., (1997) Proc. Nat'l Acad. Sci. USA 94: 11073).
Ob-R(L) has been demonstrated to transduce intracellular signaling in a manner analogous to that observed for interleukin (IL)-6 type-cytokine receptors. Ob-R(L) transmits its information via the Janus kinases (JAK), specifically Jak2 (N. Ghilardi et al., (1997) Mol. Endocrinol. 11: 393), which subsequently phosphorylate transcription factors of the STAT3 family (J. Girard (1997)).
Leptin sensitizing compounds have also been discloses. See, for example, PCT Application No. 98/02159.
3. Angiogenesis
“Angiogenesis” refers to the growth of new blood vessels, or “neovascularization,” and involves the growth of new blood vessels of relatively small caliber composed of endothelial cells. Angiogenesis is an integral part of many important biological processes including cancer cell proliferation solid tumor formation, inflammation, wound healing, repair of injured ischemic tissue, myocardial revascularization and remodeling, ovarian follicle maturation, menstrual cycle, and fetal development. New blood vessel formation is required for the development of any new tissue, whether normal or pathological, and thus represents a potential control point in regulating many disease states, as well as a therapeutic opportunity to encourage growth of normal tissue and “normal” angiogenesis.
The complete process for angiogenesis is not entirely understood, but it is known to involve the endothelial cells of the capillaries in the following ways:                (1) The attachment between the endothelial cells and the surrounding extracellular matrix (ECM) is altered, presumably mediated by proteases and glycosidases, which permit the destruction of the basement membrane surrounding the microvascular endothelial cells, thus allowing the endothelial cells to extend cytoplasmic buds in the direction of chemotacetic factors;        (2) There is a “chemotacetic process” of migration of the endothelial cells toward the tissue to be vascularized; and        (3) There is a “mitogenesis process” (e.g., proliferation of the endothelial cells to provide additional cells for new vessels).Each of these angiogenic activities can be measured independently utilizing in vitro endothelial cell cultures.        
A number of factors are known to stimulate angiogenesis. Many of these are peptide factors, and the most notable of these are the fibroblast growth factors (FGF), both acidic (aFGF) and basic (bFGF), which can be isolated from a variety of tissues including brain, pituitary and cartilage. FGFs are characterized by their heparin-binding properties. Heparin is a powerful anticoagulant agent normally found in minute amounts in the circulatory system. Other factors known to show angiogenic-stimulating activity, include but are not limited to: vascular endothelium growth factor (VEGF), angiopoietin I and II, prostaglandins E1 and E2 (B. M. Spiegelman et al., 1992), ceruloplasmin, monocyte derived monocytoangiotropin, placental angiogenic factor, glioma-derived endothelial cell growth factor, and a heparin-binding growth factor from adenocarcinoma of the kidney that is immunologically related to bFGF (R. B. Whitman et al., (1995) U.S. Pat. No. 5,470,831). Platelet-derived endothelial cell growth factor (PD-ECGF) does not stimulate proliferation of fibroblasts in contrast to the FGFs, but has demonstrated in vitro angiogenic activity (see C—H. Heldin et al., (1993) U.S. Pat. No. 5,227,302).
Factors are also known that are capable of inhibiting endothelial cell growth in vitro. One of the most extensively studied inhibitors of endothelial cell growth is protamine, which is found only in sperm. Platelet factor 4 (PF4) and major basic protein also have been demonstrated to have inhibitory effects on angiogenesis (T. Maione, (1992) U.S. Pat. No. 5,112,946). Oncostatin A, which is similar to native PF4, has also been implicated as effecting the growth of tumors and therefore may act as an angiogenesis inhibitor (T. Maione, 1992). Antibodies have also been created possessing anti-angiogenic activity (see for example, C. R. Parish (1997) U.S. Pat. No. 5,677,181). Gene therapy has also been contemplated as a means of promoting or inhibiting angiogenesis (T. J. Wickhane et al., (1996) J. Virol. 70: 6831).
4. Wound Healing and Repair of Tissue After Ischemic Injury
Wounds are internal or external bodily injuries or lesions caused by physical means, such as mechanical, chemical, bacterial, or thermal means, which disrupt the normal continuity of structures. Such bodily injuries include contusions, wounds in which the skin is unbroken, incisions, wounds in which the skin is broken by a cutting instrument, and lacerations, wounds in which the skin is broken by a dull or blunt instrument. Wounds may be caused by accidents or by surgical procedures.
Wound healing consists of a series of processes whereby injured tissue is repaired, specialized tissue is regenerated, and new tissue is reorganized. Wound healing is usually divided into three phases: the inflammatory phase, the proliferative phase, and the remodeling phase. Fibronectin has been reported to be involved in each stage of the wound healing process, particularly by creating a scaffold to which the invading cells can adhere. Initially, many mediators, such as fibronectin and fibrinogen, are released to the wound site. Thereafter, angiogenesis and re-epithelialization take place (A. Beauliu (1997) U.S. Pat. No. 5,641,483). Repair of injured tissue due to ischemia is a form of wound healing which requires extensive remodeling and re-vascularization. An infarct is, by definition, and area of tissue ischemic necrosis caused by occlusion of local blood circulation. The resulting necrotic lesion leaves the affected tissue deprived of oxygen and nutrients. In the heart, obstruction of coronary circulation in particular, results in myocardial infarction. As the ischemic myocardium undergoes rapid oxygen starvation, the hypoxic microenvironment of the infected cardiac muscle induces the synthesis of angiogenic factors to attempt re-vascularization. For example vascular endothelium growth factor (VEGF) is known to be produced in the areas of the myocardium that have undergone an infarction (Ref). Similarly, ischemic injured tissue outside the heart also produce various angiogenic factors.
Adult Wound Healing
Adult wound healing in response to injury results in restoration of tissue continuity (Adzick N. S. et al. (eds), in FETAL WOUND HEALING, Elsevier, New York 1992, Chapters 13, 12, 13 and references cited therein). While some amphibians heal by regeneration, adult mammalian tissue repair involves a complex series of biochemical events that ultimately ends in scar formation. The events occurring during wound repair resemble the process of development, including synthesis, degradation and re-synthesis of the ECM (Smith L. T. et al., (1982) J. Invest. Dermatol. 79: 935; Blanck C. E. et al., (1987) J. Cell. Biol. 105: 139(A)). The ECM contains several macromolecules, including collagen, fibronectin, fibrin, proteoglycans, and elastin. When the injury involves the dermis, repair also entails the removal of cellular debris, and the laying down of a new ECM over which epidermal continuity can be reestablished. This process of repair and dermal matrix reorganization is manifested as scar formation and maturation.
Growth Factors and Wound Healing
Manipulation of the wound healing environment by the application of extrinsic growth factors such as fibroblast growth factor (FGF) and transforming growth factors (TGFβ) (T. A. Mustoe et al., (1987) Science 237: 1333; S. M. Seyedin et al, (1986) J. Biol. Chem. 261: 5693) can influence the early stages of scar formation. During tissue repair, TGFβ modulates the inflammatory response as a potent chemoattractant for fibroblasts, macrophages, neutrophils and T lymphocytes. TGFβ can also upregulate cell surface expression of the integrins that act as receptors for fibronectin, collagen, laminin, and vitronectin thereby influencing cell adhesion and migration. TGFβ enhances the epithelial covering of exposed dermis and increases tensile strength in incision wounds. See J. W. Siebert et al., (1997) U.S. Pat. No. 5,591,716) for additional discussion of growth factors that are involved in the process of wound healing and scarring.